The direct interaction of actin with the cytoplasmic surface of the plasma membrane will be investigated by improving and modifying a new membrane isolation technique. The technique is based upon the attachment of negatively charged cells to positively charged beads. The attachment is so tenacious that attached cells can be disrupted and unattached membrane and cellular debris shorn away. The procedure will be improved by a) using reagents to neutralize sites on the beads where cells are not attached so that they do not become contaminated when cells are disrupted during membrane isolation, b) synthesizing new beads that will more firmly interact and adhere only to the outermost surface of the membrane and not cause it to distort or stretch upon attachment and c) synthesizing new beads that will gently release attached membrane for additional purification. Since the membrane on the beads is attached by its outside surface, its cytoplasmic surface is available for direct probing. The interaction of actin with this surface will be explored by determining how actin can be released from the surface and how radioactive actin can be reassociated. The nature of the reassociation coupled with cross-linking and the synthesis and use of an actin photoaffinity probe will enable an identification of the actin binding sites on the cytoplasmic surface of the plasma membrane.